Skip to main content
eScholarship
Open Access Publications from the University of California

A variety of changes, including CRISPR/Cas9-mediated deletions, in CENH3 lead to haploid induction on outcrossing.

  • Author(s): Kuppu, Sundaram
  • Ron, Mily
  • Marimuthu, Mohan PA
  • Li, Glenda
  • Huddleson, Amy
  • Siddeek, Mohamed Hisham
  • Terry, Joshua
  • Buchner, Ryan
  • Shabek, Nitzan
  • Comai, Luca
  • Britt, Anne B
  • et al.

Published Web Location

https://doi.org/10.1111/pbi.13365
Abstract

Creating true-breeding lines is a critical step in plant breeding. Novel, completely homozygous true-breeding lines can be generated by doubled haploid technology in single generation. Haploid induction through modification of the centromere-specific histone 3 variant (CENH3), including chimeric proteins, expression of non-native CENH3 and single amino acid substitutions, has been shown to induce, on outcrossing to wild type, haploid progeny possessing only the genome of the wild-type parent, in Arabidopsis thaliana. Here, we report the characterization of 31 additional EMS-inducible amino acid substitutions in CENH3 for their ability to complement a knockout in the endogenous CENH3 gene and induce haploid progeny when pollinated by the wild type. We also tested the effect of double amino acid changes, which might be generated through a second round of EMS mutagenesis. Finally, we report on the effects of CRISPR/Cas9-mediated in-frame deletions in the αN helix of the CENH3 histone fold domain. Remarkably, we found that complete deletion of the αN helix, which is conserved throughout angiosperms, results in plants which exhibit normal growth and fertility while acting as excellent haploid inducers when pollinated by wild-type pollen. Both of these technologies, CRISPR mutagenesis and EMS mutagenesis, represent non-transgenic approaches to the generation of haploid inducers.

Many UC-authored scholarly publications are freely available on this site because of the UC's open access policies. Let us know how this access is important for you.

Main Content
Current View